Beyond consensus genotyping: a case study on the Asian elephant Elephas maximus

Genetic non-invasive sampling has been a popular tool in obtaining insight into species’ ecology and biology, including demographic parameters, at a large landscape level. The possibility of inflated population estimates due to inherent genotyping errors has been established in the literature. However, the persistence of false ‘ghost’ individuals, despite widely used remedial measures, may influence population estimates and hence, requires re-examination and validation. Therefore, we used blind sub-samples of known individuals and aimed to (i) provide the extent of correct identification from faecal DNA while following standard protocols, (ii) quantify the error and (iii) suggest remedial measures using Asian elephants as a case study. We used six empirical sub-samples each from eight elephant individuals, resulting in 48 faecal DNA extracts, and genotyped across eight microsatellite markers in four replicates each to obtain consensus data. We tallied the known individual identities with the obtained genotypes of the sub-samples under several marker combinations. We observed that errors remained even after deriving consensus genotypes using a multi-tube approach. The loci-wise proportions of aberrant genotypes ranged up to 23.9%. The overall accuracy of individual assignment ranged from 18.1 to 95.2% under different marker combinations and mismatch thresholds. The combination of five markers allowing one-locus mismatch could group the highest number of empirical sub-samples (n = 28; 84.8%) correctly to their respective known genotypes. Therefore, we suggest undertaking pilot studies to screen microsatellite panels for their accuracy in assigning the identities of faecal DNA samples from known individuals using a blind test approach. Such empirical studies before undertaking large scale molecular census hold the potential to provide accurate demographic estimates.